The aims of the first study were to examine the effects of the small thiol compound pyrrolidine dithiocarbamate (PDTC) on hepatic glycogen synthesis and gluconeogenesis in type 2 diabetic rats and explore the mechanism underlying these effects. Fasting blood glucose and glycogen deposition, together with expressions of two key genes related to gluconeogenesis was studied in the liver of rats fed a normal diet (NC), high fat diet (HFD)-induced insulin resistant rats made type 2 diabetic by a single ip injection of streptozotocin (DM), and an DM with intervention of PDTC (DM+PDTC) for 1 week. The phosphorylation of Akt, GSK3&#946;and FoxO1 was assessed in liver extracts by Western blot while indirect immunofluorescence staining was performed to determine the cellular distribution of FoxO1. The DM rats exhibited obvious increase in fasting blood glucose, but a decrease in hepatic glycogen content as compared with the NC group. Activation of the Akt/GSK3&#946;pathway and subsequent inactivation of FoxO1 were greatly reduced in DM rat livers. By contrast, PDTC treatment protected DM rats against high fasting blood glucose and the loss in glycogen deposition through an increase in Akt/GSK3&#946;signaling and subsequent inactivation and nuclear export of FoxO1. Importantly, expression of phosphoenolpyruvate carboxykinase and glucose-6-phosphatase mRNAs was significantly reduced in the DM+PDTC group as compared with the DM rats. This study suggests that PDTC enhances glycogen synthesis while reducing gluconeogenesis in DM rats through Akt-dependent down-modulation of FoxO1 activity. The nutrient-dependent kinase mTOR exists in two complexes: mTOR Complex1 (mTORC1), which is rapamycin-sensitive and phosphorylates direct targets such as ribosomal protein S6 kinase (S6K1), and mTORC2, which is rapamycin-insensitive and phosphorylates Akt. The physiological roles of mTOR include involvement in metabolism, protein synthesis, apoptosis pathways, transcription factor regulation and the cell cycle. Previous studies have shown that overactivation of the mTORC1/S6K1 pathway results in inhibitory serine phosphorylation of insulin receptor substrate 1 and 2, with subsequent impairment in insulin action. Conversely, S6K1 knockout mice remain exquisitely insulin sensitive on a high-fat diet, which has been traced to the loss of a negative-feedback loop from S6K1 to the insulin receptor substrate 1. From our results, we hypothesize that PDTC improved hepatic insulin responses (increase in glycogen synthesis and reduction in gluconeogenesis) in rats made type 2 diabetic through inhibition of the mTORC1/S6K1 negative feedback loop. Studies are currently underway, using immunoprecipitation-based kinase assays and Western blots, to determine if PDTC enables recovery of the insulin receptor-insulin receptor substrate 1-PI-3 kinase pathway in various models of insulin resistance and in aging. The principal aim of the second study was to perform a global gene expression profiling of human liver-derived HepG2 cells after treatment with PDTC or IL-6 for up to 8 h. We have reported earlier that PDTC antagonizes the cellular responsiveness to IL-6 through impairment in STAT3 activation and downstream signaling. Here, through an unbiased pathway analysis method, gene array analysis showed dramatic and temporal differences in expression changes in response to PDTC vs. IL-6. A significant number of genes associated with metabolic pathways, inflammation, translation and mitochondrial function were changed, with ribosomal protein genes and DDIT4 primarily up-regulated with PDTC but down-regulated with IL-6. Quantitative PCR and Western blot analyses validated the microarray data and showed the reciprocal expression pattern of the mTOR negative regulator, DDIT4, in response to PDTC vs. IL-6. Administration of PDTC resulted in a rapid and sustained activation of Akt, and subsequently blocked IL-6-mediated increase in mTOR complex 1 (mTORC1) function through upregulation in DDIT4 expression. Conversely, the decline in IL-6-dependent mTOR activation by PDTC was slowed down in DDIT4-knockdown cells. The overall protein biosynthetic capacity of the cells was severely blunted by IL-6 but increased in a rapamycin-independent pathway by PDTC. These results demonstrate a critical effect of PDTC on mTORC1 function and provide evidence that PDTC can reverse IL-6-related signaling via induction of DDIT4. Elevated circulating levels of IL-6 is one of many factors involves in the pathophysiology of chronic inflammatory diseases. It is interesting that IL-6 is considered to be a proteolysis-inducing factor due to its modulation of muscle proteolytic systems and that the muscle wasting observed in aged, sarcopenic rats is associated with enhanced activity of the proteasome pathway and reduced synthesis of muscle proteins. Both preclinical and clinical studies have recently shown that PDTC and derivatives target the ubiquitin-proteasome system, which might explain the protection offered by PDTC against muscle and adipose tissue loss in a mouse model of cachexia. Recent findings indicate that suppression of the ubiquitin-proteasome system induces inhibition of mTORC1 signaling. We hypothesize that the ability of PDTC to inhibit mTORC1 stems from proteasomal stabilization of DDIT4, the short-lived negative repressor of mTORC1. These findings suggest that cellular sensitivity to proteasome inhibition and mTORC2/Akt pathway activation by PDTC may represent a novel strategy to enhance skeletal muscle satellite cell proliferation and differentiation.